Genetica 115: 37–47, 2002. 37 © 2002 Kluwer Academic Publishers. Printed in the Netherlands. Feast and famine in plant genomes Jonathan F. Wendel1, Richard C. Cronn1, J. Spencer Johnston2 & H. James Price3 1Department of Botany, Iowa State University, Ames, IA, 50011, USA (Phone: +1-515-294-7172; Fax: +515­ 294-1337; E-mail: [email protected]); 2Department of Entomology, 3Department of Soil and Crop Sciences, Texas A & M University, College Station, TX 77843, USA Key words: cotton, DNA content, genome evolution, genome size, Gossypium, Malvaceae, molecular evolution, 2C values Abstract Plant genomes vary over several orders of magnitude in size, even among closely related species, yet the origin, genesis and significance of this variation are not clear. Because DNA content varies over a sevenfold range among diploid species in the cotton genus (Gossypium) and its allies, this group offers opportunities for exploring patterns and mechanisms of genome size evolution. For example, the question has been raised whether plant genomes have a ‘one-way ticket to genomic obesity’, as a consequence of retroelement accumulation. Few empirical studies directly address this possibility, although it is consistent with recent insights gleaned from evolutionary genomic investigations. We used a phylogenetic approach to evaluate the directionality of genome size evolution among Gos­ sypium species and their relatives in the cotton tribe (Gossypieae, Malvaceae). Our results suggest that both DNA content increase and decrease have occurred repeatedly during evolution. In contrast to a model of unidirectional genome size change, the frequency of inferred genome size contraction exceeded that of expansion. In conjunction with other evidence, this finding highlights the dynamic nature of plant genome size evolution, and suggests that poorly understood genomic contraction mechanisms operate on a more extensive scale that previously recognized. Moreover, the research sets the stage for fine-scale analysis of the evolutionary dynamics and directionality of change for the full spectrum of genomic constituents. Introduction massive accumulation of retroelements, as elegantly shown for some species in the grass family (Bennet­ Plant genomes exhibit extraordinary variation in size, zen, 1996; SanMiguel et al., 1996; Bennetzen, 2000). ranging from approximately 125 Mbp in Arabidopsis The latter observations led naturally to a proposal that thaliana, known with exquisite precision now that genome size evolution in plants may largely be uni­ the genome has been nearly completely sequenced directional, from small to large (Bennetzen & Kellogg, (The Arabidopsis Genome Initiative, 2000), to over 1997a), with an overall pattern of ratcheting upward 120,000 Mbp in some relatives of lilies (Bennett & due to the combined effects of polyploidization and Leitch, 1995; Bennett et al., 1997; Leitch et al., 1998). retroelement accumulation. Because this remarkable range in DNA content is not The suggestion that plants may have a ‘one-way associated with variation in the basic complement of ticket to genomic obesity’ (Bennetzen & Kellogg, genes required for growth and development, it has 1997a) provided an appealing scenario that appeared become renowned as the ‘C-value paradox’ (Thomas, to account, at least in part, for the C-value paradox. 1971). Mechanistic explanations for the observed size Plants with small genomes are suggested to have been variation include repeated cycles of polyploidy over relatively effective in suppressing retrotransposon evolutionary time (Leitch & Bennett, 1997; Soltis & activity; whereas those with large genomes achieved Soltis, 1999; Otto & Whitton, 2000; Wendel, 2000), this condition by release from suppression (or new leading to progressively larger genomes, as well as invasion) and massive retroelement amplification. Be­ 38 cause the generality of this proposal is unknown the tribe using exemplar sampling, filled in gaps in and because alternative explanations exist, the sub­ our knowledge of chromosome numbers to ensure that ject has attracted considerable attention (Bennetzen & only diploids were included in the study, and em­ Kellogg, 1997b; Petrov, 1997; Leitch et al., 1998; ployed analytical methods for inferring ancestral con­ Vicient et al., 1999; Federoff, 2000; Rabinowicz, ditions for quantitative varying characters (Maddison, 2000; Shirasu et al., 2000; Petrov, 2001). The hy­ 1991; Martins & Hansen, 1997; Cunningham et al., pothesis of unidirectional genome size change has 1998; Martins, 1999; Oakley & Cunningham, 2000). been rigorously tested in only a few cases (Bennetzen & Kellogg, 1997a; Cox et al., 1998; Leitch et al., Materials and methods 1998). In addition, accumulating evidence suggests that genome size contraction may also be a com­ Organismal context and taxon sampling mon evolutionary occurrence. Supporting this sugges­ tion is recent evidence that underscores the diversity Included in the eight genera of the Gossypieae are four and scope of genomic deletional mechanisms (Petrov small genera with restricted geographic distributions et al., 1996; Petrov, 1997; Vicient et al., 1999; Kirik (Fryxell, 1979). Lebronnecia Fosberg, from the Mar­ et al., 2000; Petrov et al., 2000; Shirasu et al., 2000; quesas Islands, is a monotypic genus, as is Cephalo­ Petrov, 2001), the phylogenetically widespread dis­ hibiscus Ulbrich, from New Guinea and the Solomon tribution of plants with small genomes (Bennett & Islands. Gossypioides Skovsted ex J.B. Hutchinson Leitch, 1995; Bennett et al., 1997; Leitch et al., 1998), contains two species from East Africa and Madagas­ and the prevalence of high levels of infraspecific DNA car. Kokia Lewton, endemic to the Hawaiian Islands, content variation (Price, 1988; Kalendar et al., 2000) includes three extant and one extinct species. In ad­ that are not easily reconciled with an assumption of dition to these four small genera, the tribe includes unidirectional evolutionary change. four moderately sized genera with broader geographic A prerequisite for any discussion of the direction­ ranges: Hampea Schlechtendal comprises 21 neo­ ality of genome size change is that the phylogenetic tropical species; Cienfuegosia Cavanilles includes 25 relationships of the organisms under study are un­ species with an aggregate range that includes the neo­ derstood. Phylogenetic analysis provides an essential tropics and parts of Africa; 17 species are recognized framework for inferences of ancestral genome sizes in the pantropical genus Thespesia Solander; and Gos­ (Bennetzen & Kellogg, 1997a) as well as for polar­ sypium, as noted above, is the largest genus in the tribe izing genomic changes for specific homologous gen­ with about 50 species. omic regions. Hence, robust phylogenies are essential An initial phylogenetic scaffolding for the tribe for both global (genome size) and local (fine-scale) was erected several years ago based on sequences inferences of past genomic evolutionary events. A from the chloroplast gene ndhF and from nuclear particularly useful model in this respect is the group ribosomal ITS sequences (Seelanan et al., 1997). of plants comprising the cotton genus (Gossypium These analyses provided unequivocal support for L.) and its relatives in the small Malvaceous tribe several relationships among genera but left other as­ Gossypieae Alefeld. This monophyletic (LaDuke & pects of their evolutionary history unclear. Partic­ Doebley, 1995) tribe includes eight genera (Fryxell, ularly strongly supported was the close relationship 1979), the largest being Gossypium with approxim­ between Gossypioides and Kokia and their collec­ ately 50 species (Fryxell, 1992). Remarkable genome tive sister-taxon relationship to Gossypium. Thespesia size variation exists among the 45 diploid species of was implicated to be at least biphyletic and perhaps Gossypium, with 2C DNA contents ranging from ap­ polyphyletic, with a portion being sister to Lebron­ proximately 2.0 to 7.0 pg (Wendel et al., 1999). This necia and perhaps Hampea and the remainder be­ information motivated an exploration of genome sizes ing cladistically unresolved along with Cienfuegosia among its relatives, as well as phylogenetic relation­ and the (Hampea + Lebronnecia + Thespesia- in part) ships among the genera in the tribe. Our intention clade. Cephalohibiscus was not included in these was to generate the information required to understand analyses as it was not available. This earlier work the genome size variation that exists among diploid provided some insights into the evolutionary history species within this single tribe, and thereby facilitate of the group, but left unanswered several key questions inferences of the directionality of genome size change. of branching order, particularly as regards the earliest To accomplish this we estimated the phylogeny of divergences within the tribe. 39 Table 1. Species in the Gossypieae used for phylogenetic analysis, chromosome counts, and genome size measurementsa Taxon Chromosome Genome no. (2n) size(2C) C. hitchcockii (Ulbrich ex Kearney) Blanchard 20 2.3 C. tripartita H.B.K. Gürke 20 1.9 C. yucatanensis Millspaugh 22 2.0 G. kirkii (Mast.) J.B. Hutchinson 24 1.2 G. herbaceum L. 26 3.7 G. raimondii Ulbrich 26 2.0 H. appendiculata (J. Donnell-Smith) Standley 26 5.9 K. drynarioides (Seemann) Lewton 24 1.2 L. kokioides Fosberg 26 3.6 T. lampas (Cavanilles) Dalzell ex Dalzell and Gibson 26 3.2 T. populnea (L.) Solander ex Correa 26 8.2 T. thespesioides (R. Brown ex Bentham) Fryxell 26 3.2 M. sylvestris